Method for Treating a Pulmonary Disease State in Mammals by Up Regulating Indigenous in vivo Levels of Inflammatory Agents in Mammalian Cells

ABSTRACT

The present invention provides novel methods for treating a pulmonary disease state in mammals by up-regulating indigenous in vivo levels of an inflammatory agent in mammalian cells comprising contacting the mammalian cells with a therapeutically effective amount of an inflammatory regulator and a pharmaceutical agent. The inflammatory agent is selected from the group consisting of cytokines, transforming growth factor-β, elastase, and white blood cells, and wherein the inflammatory regulator is selected from the group consisting of pyruvates and pyruvate precursors. The pharmaceutical agent is selected from the group comprising anti-bacterial agents, anti-virals, anti-fungals, anti-tumors, antihistamines, proteins, enzymes, hormones such as insulin, non-steroidal anti-inflammatories, cytokines, steroids, and nicotine.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is a national phase entry under 35 U.S.C. §371of International Application No. PCT/US2012/042254 filed Jun. 13, 2012,which claims the benefit of priority of a continuation-in-partapplication Ser. No. 12/009,649 filed 22 Jan. 2008 which is acontinuation-in-part application of U.S. patent application Ser. No.11/890,911, filed 8 Aug. 2007 now U.S. Pat. No. 8,114,907 to Martin, thedisclosures of which are incorporated herein by reference.

FIELD OF THE INVENTION

The present invention provides novel methods for treating a pulmonarydisease state in mammals by up regulating indigenous in vivo levels ofan inflammatory agent in mammalian cells comprising contacting themammalian cells with a therapeutically effective amount of aninflammatory regulator, wherein the inflammatory agent is selected fromthe group consisting of cytokines, transforming growth factor-β,elastase, and white blood cells, and wherein the inflammatory regulatoris selected from the group consisting of pyruvates and pyruvateprecursors.

BACKGROUND OF THE INVENTION

The disclosures referred to herein to illustrate the background of theinvention and to provide additional detail with respect to its practiceare incorporated herein by reference and, for convenience, arereferenced in the following text and respectively grouped in theappended bibliography.

Inflammatory agents are produced by a wide variety of body cells and arenatural proteins produced by the cells of the immune system of mostvertebrates in response to challenges by foreign agents such as viruses,bacteria, parasites, and tumor cells (1). There are many physiologicaldisease states in man wherein which the human body's immune systemproduces abnormally low levels of inflammatory agents. In theseconditions, the body needs an inflammatory regulator to increase orup-regulate specific inflammatory agents to fight infections in thelungs and sinuses and/or to enhance a balanced immune system. Diseasesof the human body that results in the production of abnormally lowinflammatory agents including IL-10, include HIV, Humoral ImmuneDeficiency, Alzheimer's, Interstitial lung disease, Sarcoidosis, CysticFibrosis, T-cell deficiency, Neutropenia, Asplenia, complementdeficiency, allergic Rhinitis, sinusitis, COPD, Asthma. All of thesediseases result in the body incurring an imbalance to the immune systemthat leads to high rates of infections, including infections in thesinuses and lungs.

There are also medications and drugs that once administered, lower thebody's own immune systems inflammatory agents which results in theeventual onset of infections and injury to normal cells and membranes.These drugs include steroids, cancer drugs, alkylating agents,anti-metabolites, Azathioprine and mercaptopurine (immunosuppressiveagents) antibiotics, antibodies, cyclosporine, sirolimus, nicotine, andinsulin. Other irritants like chemicals, irritant gases such as inhaleddiesel fumes and cigarette smoke also lower certain inflammatory agentsof the immune system needed to maintain a healthy immune system in thelungs and sinuses.

When inflammation is activated, many inflammatory cytokines becomeelevated, while other components of the immune system become suppressed,causing an imbalance in the immune system, that can injure cells andorgans. The present invention comprises compositions and methods thatbring balance back to the immune system. When certain inflammatoryagents are abnormally high, the levels of other inflammatory agents arereduced or shut down. It is at this point, that the use of aninflammatory regulator is appropriate to up-regulate these reducedinflammatory agents and bring a necessary balance back to the immunesystem, allowing it to properly fight infections and prevent tissuedamage. It has been been surprisingly discovered that high levels ofinhaled pyruvate enhance the up-regulation of depressed levels ofcertain cytokines that have been lowered by the increased production ofothers so as to balance the negative effects of the other elevatedcytokines and allows the immune system to function properly to fightinfections and decrease the likelihood of tissue damage that may resultfrom the swelling and fluid imbalance in cells. For example, IL-10 iscapable of inhibiting the synthesis of pro-inflammatory cytokines andalso displays a potent ability to suppress an antigen induced reaction.Therefore, up-regulating IL-10 has been shown to be effective intreating allergic rhinitis in the following examples.

Cytokines are a group of proteins and peptides that are used inorganisms as signaling compounds and are used to allow one cell tocommunicate with another. The cytokine family consists mainly of smallerwater-soluble proteins and glycoproteins. Cytokines are released by manytypes of cells, principally activated lymphocytes, and macrophages butalso endothelium, epithelium and connective tissue. They areparticularly important in both innate and adaptive immune responses. Dueto their central role in the immune system, cytokines are involved in avariety of immunological, inflammatory and infectious diseases.

Interleukins (Ls) are a group of inflammatory cytokines that were firstseen to be expressed by white blood cells. Interleukins are produced bya wide variety of bodily cells including endothelial cells andmacrophages. The family of interleukins includes IL-1 to IL-33. Thefunction of the immune system depends in a large part on interleukins,and rare deficiencies of a number of them have been described, allfeaturing autoimmune diseases or immune deficiency.

Interferons (IFNs) belong to a large class of glycoproteins and arecytokines. Interferons are natural proteins produced by the cells of theimmune system of most vertebrates in response to challenges by foreignagents such as viruses, bacteria, parasites and tumor cells. Interferonsassist the immune response by inhibiting viral replication within othercells of the body.

Tumor necrosis factor (TNF) is a cytokine involved in systemicinflammation and is a member of a group of cytokines that stimulate theacute phase reaction. Tumor necrosis factor causes apoptotic cell death,cellular proliferation, differentiation, inflammation, tumorigenesis,and viral replication. Tumor necrosis factors primary role is in theregulation of immune cells. Dysregulation and, in particular,overproduction of tumor necrosis factor have been implicated in avariety of human diseases, as well as cancer

Chemokines are a family of small cytokines, or proteins that areclassified according to shared structural characteristics such as smallsize (they are all approximately 8-10 kilo Daltons in size), and thepresence of four cysteine residues in conserved locations that are keyto forming their 3-dimensional shape. Chemokines have the ability toinduce directed chemotaxis in nearby responsive cells (chemotacticcytokines). Some chemokines are considered pro-inflammatory and can beinduced during an immune response to promote cells of the immune systemto a site of infection, while others are considered homeostatic and areinvolved in controlling the migration of cells during normal processesof tissue maintenance or development. Chemokines exert their biologicaleffects by interacting with G protein-linked transmembrane receptorscalled chemokine receptors that are selectively found on the surfaces oftheir target cells.

Cytokines and chemokines have the ability to stimulate leukocytemovement and play an important role in inflammation. Cytokines caninfluence the synthesis of other cytokines and chemokines. Cytokines canalso stimulate cell proliferation acting as growth factors. Cytokinesthat regulate lymphocyte activation, growth and differentiation includeIL-2, IL-4, IL-10, and TNF-β. Cytokines involved with natural immunity,inflammation, include TNF-α, IL-1, INF-α, INF-β, and IL-6. Cytokinesthat activate inflammatory cells like macrophages include IFN-γ, TNF-α,TNF-β, IL-5, IL-10, IL-12, and IL-8. Cytokines that stimulatehemopoiesis and mediate immature leukocyte growth and differentiationinclude IL-3, IL-7, c-kit ligand, granulocyte-macrophage, granulocytecolony-stimulating factor (G-CSF), and stem cell factor. Granulocytecolony-stimulating factor is a glycoprotein, growth factor or cytokineproduced by a number of different tissues to stimulate the bone marrowto produce granulocytes and stem cells. Granulocyte colony-stimulatingfactor then stimulates the bone marrow to pulse them out of the marrowinto the blood.

IL-8 is responsible in attracting white blood cells to the site ofinfection. The major cytokines that mediate inflammation are IL-1, IL-8,and TNF (α and β). IL-1 and TNF-α are produced by activated macrophages.Their secretion can be stimulated by infections, endotoxins, immunecomplexes, toxins, physical injury, and a variety of inflammatoryprocesses. Their most important actions in inflammation are their effecton endothelium, leukocytes, and fibroblasts and induction of thesystemic acute phase reactions. TNF also cause aggregation and primingof neutrophils, leading to a release of proteolytic enzymes, thuscontributing to tissue damage. TNF-α, IL-1, and IL-6 also induce thesystemic acute phase responses associated with infection, or injury,including fever, loss of appetite, the production of slow wave sleep,release of neutrophils into circulation, release of hormones,hemodynamic effects of septic shock, hypotension, decrease in vascularresistance, increased heart rate, and decrease in blood pH.

An excess of inflammatory agents can increase the production of oxygenradicals, including superoxide anions and hydrogen peroxide, producedduring the inflammatory phase of an injury, which will destroy healthytissue surrounding the site and will mitigate the positivebronchodilation effect of nitric oxide (26). Oxygen radicals can alsoinitiate lipid peroxidation employing arachidonic acid as a substrateproducing prostaglandins and leukotrienes. Hydrogen peroxide (H₂O₂) caninduce arachidonic acid metabolism in alveolar macrophages. Oxygenradicals also produce 8-isoprostanes, which are potent renal andpulmonary artery vasoconstrictors, bronchoconstrictors, and induceairflow obstructions (26, 27). Because oxygen radicals contribute to theinstability of nitric oxide, the addition of superoxide dismutase (SOD)or catalase (15) or Vitamin E (28) protect nitric oxide to produce itsdesired bronchodilation (2). Hydrogen peroxide is elevated in patientswith chronic obstructive pulmonary disease (COPD), asthma, and AcuteRespiratory Distress Syndrome (ARDS) (26). A study in 28 patients showeda significant correlation between oxygen radical generation in whiteblood cell count (WBC) and the degree of bronchial hyperreactivity invivo in non-allergic patients (18). Thus the ability of pyruvate toregulate inflammation, and inflammatory agents, which can increase thesynthesis of oxygen radicals, should reduce the production of oxygenradicals when needed.

Sodium pyruvate is an antioxidant that reacts directly with oxygenradicals to neutralize them. In macrophages, and other cell lines,sodium pyruvate regulates the level of oxygen radicals by acting as anantioxidant and also increases the synthesis of nitric oxide (9). It canspecifically lower the overproduction of superoxide anions. Sodiumpyruvate also increases cellular levels of glutathione, a major cellularantioxidant (12). It was recently discovered that glutathione is reduceddramatically in antigen-induced asthmatic patients (13) and inhaledglutathione does not readily enter cells. Pyruvate does enter all cellsvia a transport system and can also cross the blood brain barrier.Excess sodium pyruvate beyond that needed to neutralize oxygen radicalswill enter the bronchial and lung cells. All cells have a transportsystem that allow cells to concentrate pyruvate at higher concentrationsthan serum levels. In the cell, pyruvate raises the pH level, increaseslevels of ATP, decreasing levels of ADP and cAMP, and increases levelsof GTP, while decreasing levels of cGMP. Nitric oxide (NO) acts in theopposite mode by increasing levels of cGMP and ADP, and requires anacidic pH range in which to work (19). While the above therapeuticcompositions and methods are reported to inhibit the production ofreactive oxygen intermediates, like hydrogen peroxide or peroxynitrite,none of the disclosures describe methods for treating a pulmonarydisease state in mammals by regulating indigenous in vivo levels ofinflammatory agents in mammalian cells.

U.S. Pat. No. 6,063,407 (Zapol et al.) discloses methods of treating,inhibiting or preventing vascular thrombosis or arterial restenosis in amammal. The methods include causing the mammal to inhale atherapeutically effective concentration of gaseous nitric oxide. Theinhaled nitric oxide may further comprise compounds that potentiate thebeneficial effects of inhaled nitric oxide and antithrombotic agentsthat complement or supplement the beneficial effects of inhaled nitricoxide.

U.S. Pat. No. 6,020,308 (Dewhirst et al.) discloses the use of aninhibitor of nitric oxide activity, such as a nitric oxide scavenger ora nitric oxide synthase inhibitor, as an adjunct to treatment ofinappropriate tissue vascularization disorders.

U.S. Pat. No. 5,891,459 (Cooke et al.) discloses the maintenance orimprovement of vascular function and structure by long termadministration of physiologically acceptable compounds, such asL-arginine, L-lysine, physiologically acceptable salts thereof, andpolypeptide precursors thereof, which enhance the level of endogenousnitric oxide or other intermediates in the nitric oxide inducedrelaxation pathway in the host. The method further comprises theadministration of other compounds, such as B6, folate, B12, or anantioxidant, which provide for short-term enhancement of nitric oxide.

U.S. Pat. No. 5,873,359 (Zapol et al.) discloses a method for treatingor preventing bronchoconstriction or reversible pulmonaryvasoconstriction in a mammal, which method includes causing the mammalto inhale a therapeutically effective concentration of gaseous nitricoxide or a therapeutically effective amount of a nitric oxide releasingcompound and an inhaler device containing nitric oxide gas and/or anitric oxide releasing compound.

U.S. Pat. No. 5,767,160 (Kaesemeyer) discloses a therapeutic mixturecomprising L-arginine and an agonist of nitric oxide synthase, such asnitroglycerin for the treatment of diseases related to vasoconstriction.The vasoconstriction is relieved by stimulating the constitutive form ofnitric oxide synthase (cNOS) to produce native nitric oxide. The nativenitric oxide has superior beneficial effect when compared to exogenousnitric oxide produced by an L-arginine independent pathway in terms ofthe ability to reduce clinical endpoints and mortality.

U.S. Pat. No. 5,543,430 (Kaesemeyer) discloses a therapeutic mixturecomprising a mixture of L-arginine and an agonist of nitric oxidesynthase such as nitroglycerin for the treatment of diseases related tovasoconstriction. The vasoconstriction is relieved by stimulating theconstitutive form of nitric oxide synthase to produce native nitricoxide. The native nitric oxide has superior beneficial effect whencompared to exogenous nitric oxide produced by an L-arginine independentpathway in terms of the ability to reduce clinical endpoints andmortality.

U.S. Pat. No. 5,428,070 (Cooke et al.) discloses a method for treatingatherogenesis and restenosis by long-term administration ofphysiologically acceptable compounds, which enhance the level ofendogenous nitric oxide in the host. Alternatively, or in combination,other compounds may be administered which provide for short-termenhancement of nitric oxide, either directly or by physiologicalprocesses. In addition, cells may be genetically engineered to provide acomponent in the synthetic pathway to nitric oxide, so as drive theprocess to enhance nitric oxide concentration, particularly inconjunction with the administration of a nitric oxide precursor.

U.S. Pat. No. 5,286,739 (Kilbourn et al.) discloses an anti-hypotensiveformulation comprising a mixture of amino acids, which is essentiallyarginine free or low in arginine (less than about 0.1%, most preferably,about 0.01%). The formulation may include ornithine, citrulline, orboth. A method for prophylaxis and treatment of systemic hypotension inan animal is also provided. A method for treating hypotension caused bynitric oxide synthesis through administering a low or essentiallyarginine free parenteral formulation to an animal, so as to reduce oreliminate nitric oxide synthesis is described. A method for treating ananimal in septic shock is also disclosed, comprising administering tothe animal an anti-hypotensive formulation comprising a mixture of aminoacids, which is essentially arginine free. Prophylaxis or treatment ofsystemic hypotension, particularly that hypotension incident tochemotherapeutic treatment with biologic response modifiers, such astumor necrosis factor or interleukin-1 or 2, may be accomplished throughthe administration of the defined anti-hypotensive formulations untilphysiologically acceptable systolic blood pressure levels are achievedin the animal. Treatment of an animal for septic shock induced byendotoxin may also be accomplished by administering to the animal thearginine free formulations.

U.S. Pat. No. 5,217,997 (Levere et al.) discloses a method for treatinga high vascular resistance disorder in a mammal by administering to amammalian organism in need of such treatment a sufficient amount ofL-arginine or pharmaceutically acceptable salt thereof to treat a highvascular resistance disorder. The L-arginine is typically administeredin the range of about 1 mg to 1500 mg per day. High vascular resistancedisorders include hypertension, primary or secondary vasospasm, anginapectoris, cerebral ischemia and preeclampsia. Also disclosed is a methodfor preventing or treating bronchial asthma in a mammal by administeringto a mammalian organism in need of such prevention or treatment asufficient amount of L-arginine to prevent or treat bronchial asthma.

U.S. Pat. No. 5,158,883 (Griffith) discloses pharmaceutically purephysiologically active NG-aminoarginine (i.e., the L or D, L form), orpharmaceutically acceptable salts thereof, administered in a nitricoxide synthesis inhibiting amount to a subject in need of suchinhibition (e.g., a subject with low blood pressure or needingimmunosuppressive effect) or added to a medium containing isolatedorgans, intact cells, cell homogenates or tissue homogenates in anamount sufficient to inhibit nitric oxide formation to elucidate orcontrol the biosynthesis, metabolism or physiological role of nitricoxide.

U.S. Pat. Nos. 5,798,388, 5,939,459, and 5,952,384 (Katz) pertain tomethods for treating various disease states in mammals caused bymammalian cells involved in the inflammatory response and compositionsuseful in the method. The method comprises contacting the mammaliancells participating in the inflammatory response with an inflammatorymediator. The inflammatory mediator is present in an amount capable ofreducing the undesired inflammatory response and is an antioxidant. Thepreferred inflammatory mediator is a pyruvate. Katz discloses thetreatment of airway diseases of the lungs such as bronchial asthma,acute bronchitis, emphysema, chronic obstructive emphysema,centrilobular emphysema, panacinar emphysema, chronic obstructivebronchitis, reactive airway disease, cystic fibrosis, bronchiectasis,acquired bronchiectasis, kartaagener's syndrome; atelectasis, acuteatelectasis, chronic atelectasis, pneumonia, essential thrombocytopenia,legionnaires disease, psittacosis, fibrogenic dust disease, diseases dueto organic dust, diseases due to irritant gases and chemicals,hypersensitivity diseases of the lung, idiopathic infiltrative diseasesof the lungs and the like by inhaling pyruvate containing compositions.

U.S. Pat. No. 5,296,370 (Martin et al.) discloses therapeuticcompositions for preventing and reducing injury to mammalian cells andincreasing the resuscitation rate of injured mammalian cells. Thetherapeutic composition comprises (a) pyruvate selected from the groupconsisting of pyruvic acid, pharmaceutically acceptable salts of pyruvicacid, and mixtures thereof, (b) an antioxidant, and (c) a mixture ofsaturated and unsaturated fatty acids wherein the fatty acids are thosefatty acids required for the resuscitation of injured mammalian cells.

U.S. Pat. No. 6,689,810 (Martin) discloses a therapeutic composition fortreating pulmonary diseases states in mammals by altering indigenous invivo levels of nitric oxide. The therapeutic composition consists ofpyruvates, pyruvate precursors, α-keto acids having four or more carbonatoms, precursors of α-keto acids having four or more carbons, and thesalts thereof.

U.S. Pat. No. 7,122,578 (Martin) discloses a therapeutic composition fortreating topical diseases states and injuries in mammals involvinginjuries, which cause pain, erythema, swelling, crusting, ischemia,scarring, and excess white blood cell infiltration. The method involvesthe use of α-keto acids to suppress inflammation.

WO 2006/086643 (Martin) discloses a non-pulmonary treatment of mammaliandiseases and injuries caused by the over-expression of peroxynitrite.

While the above therapeutic compositions and methods are reported toinhibit the production of reactive oxygen intermediates, such ashydrogen peroxide, peroxynitrite or nitric oxide, none of thedisclosures describe a method for treating a pulmonary disease state inmammals by altering indigenous in vivo levels of inflammatory agents.

SUMMARY OF THE INVENTION

The present invention provides novel methods for treating a pulmonarydisease state in mammals by up regulating indigenous in vivo levels ofan inflammatory agent in mammalian cells comprising contacting themammalian cells with a therapeutically effective amount of aninflammatory regulator, wherein the inflammatory agent is selected fromthe group consisting of cytokines, transforming growth factor-β,elastase, and white blood cells, and wherein the inflammatory regulatoris selected from the group consisting of pyruvates and pyruvateprecursors.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph illustrating individual sputum total protein levelsbefore and after study drug inhalation. Slash marks represent the medianlevel.

FIG. 2 is a graph illustrating median change from pre- to post-studydrug inhalation in sputum total protein levels and by drug dose level.

FIG. 3 is a graph illustrating individual sputum free elastase levelsbefore and after study drug inhalation. Slash marks represent the medianlevel.

FIG. 4 is a graph illustrating median change from pre- to post-studydrug inhalation in sputum free elastase levels and by drug dose level.

FIG. 5 is a graph illustrating individual sputum IL-6 levels before andafter study drug inhalation. Slash marks represent the median level.

FIG. 6 is a graph illustrating median change from pre- to post-studydrug inhalation in sputum IL-6 levels and by drug dose level.

FIG. 7 is a graph illustrating individual sputum IL-8 levels before andafter study drug inhalation. Slash marks represent the median level.

FIG. 8 is a graph illustrating median change from pre- to post-studydrug inhalation in sputum IL-8 levels and by drug dose level.

FIG. 9 is a graph illustrating individual sputum TNF-α level before andafter study drug inhalation. Slash marks represent the median level.

FIG. 10 is a graph illustrating median change from pre- to post-studydrug inhalation in sputum TNF-α levels and by drug dose level.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides novel methods for treating a pulmonarydisease state in mammals by up regulating indigenous in vivo levels ofan inflammatory agent in mammalian cells comprising contacting themammalian cells with a therapeutically effective amount of aninflammatory regulator, wherein the inflammatory agent is selected fromthe group consisting of cytokines, transforming growth factor-β,elastase, and white blood cells, and wherein the inflammatory regulatoris selected from the group consisting of pyruvates and pyruvateprecursors.

As used herein, the following terms have the given meanings:

The term “injured cell” as used herein refers to a cell which has someor all of the following: (a) injured membranes so that transport throughthe membranes is diminished and may result in one or more of thefollowing, an increase in toxins and normal cellular wastes inside thecell and/or a decrease in nutrients and other components necessary forcellular repair inside the cell, (b) an increase in concentration ofoxygen radicals inside the cell because of the decreased ability of thecell to produce antioxidants and enzymes, and (c) damaged DNA, RNA andribosomes which must be repaired or replaced before normal cellularfunctions can be resumed.

The term “pharmaceutically acceptable,” such as pharmaceuticallyacceptable carriers, excipients, etc., refers to pharmacologicallyacceptable and substantially non-toxic to the subject to which theparticular compound is administered.

The term “pharmaceutically acceptable salt” refers to conventionalacid-addition salts or base-addition salts that retain the biologicaleffectiveness and properties of the compounds of the present inventionand are formed from suitable non-toxic organic or inorganic acids ororganic or inorganic bases. Sample acid-addition salts include thosederived from inorganic acids such as hydrochloric acid, hydrobromicacid, hydroiodic acid, sulfuric acid, sulfamic acid, phosphoric acid andnitric acid, and those derived from organic acids such asp-toluenesulfonic acid, salicylic acid, methanesulfonic acid, oxalicacid, succinic acid, citric acid, malic acid, lactic acid, fumaric acid,and the like. Sample base-addition salts include those derived fromammonium, potassium, sodium, and quaternary ammonium hydroxides, such asfor example, tetramethylammonium hydroxide. Chemical modification of apharmaceutical compound (i.e., drug) into a salt is a technique wellknown to pharmaceutical chemists to obtain improved physical andchemical stability, hydroscopicity, and solubility of compounds. See,e.g., H. Ansel et. al., Pharmaceutical Dosage Forms and Drug DeliverySystems (6^(th) Ed. 1995) at pp. 196 and 1456-1457.

The term “prodrug” or “precursor” refers to compounds, which undergobiotransformation prior to exhibiting their pharmacological effects. Thechemical modification of drugs to overcome pharmaceutical problems hasalso been termed “drug latentiation.” Drug latentiation is the chemicalmodification of a biologically active compound to form a new compound,which upon in vivo enzymatic attack will liberate the parent compound.The chemical alterations of the parent compound are such that the changein physicochemical properties will affect the absorption, distributionand enzymatic metabolism. The definition of drug latentiation has alsobeen extended to include nonenzymatic regeneration of the parentcompound. Regeneration takes place as a consequence of hydrolytic,dissociative, and other reactions not necessarily enzyme mediated. Theterms prodrugs, latentiated drugs, and bio-reversible derivatives areused interchangeably. By inference, latentiation implies a time lagelement or time component involved in regenerating the bioactive parentmolecule in vivo. The term prodrug is general in that it includeslatentiated drug derivatives as well as those substances, which areconverted after administration to the actual substance, which combineswith receptors. The term prodrug is a generic term for agents, whichundergo biotransformation prior to exhibiting their pharmacologicalactions.

The term “therapeutically effective amount” refers to an amount of atherapeutically effective compound, or a pharmaceutically acceptablesalt thereof, which is effective to treat, prevent, alleviate orameliorate symptoms of a disease.

Pyruvates can act as inflammatory mediators (antioxidants) to neutralizeoxygen radicals directly, thus lowering the level of inflammation.Pyruvates can also act as antioxidants to regulate the synthesis ofnitric oxide. The regulation of oxygen radicals and the synthesis ofnitric oxide operate by a different set of genes than those thatregulate the synthesis of inflammatory agents such as cytokines and thusoperates by a different mechanism. Applicant has discovered thatpyruvates and pyruvate precursors can up or down regulate indigenous invivo levels of inflammatory agents such as cytokines to regulateinflammation. Specifically, applicant has discovered that pyruvates inlow dosage amounts, can down regulate the production of inflammatoryagents and the number of white blood cells to stop the negative sideeffects of chronic inflammation in uninfected pulmonary diseases or, inhigh dosage amounts, can up-regulate the production of inflammatoryagents and the number of white blood cells needed to kill infections orcancer in infected pulmonary diseases. Mediation of inflammation is verydifferent from the regulation of inflammation. Mediation is a directchemical effect on the inflammatory components such as the ability ofpyruvates to act as antioxidants against oxygen radicals such ashydrogen peroxide, peroxynitrite, or nitric oxide to elicit a response.Regulation of inflammation, such as the up or down regulation of thelevels of inflammatory agents, is a direct effect of pyruvates to elicita response on a genetic level and to specifically effect and regulatethe function of inflammatory cells such as white blood cells. Theability to regulate cellular functions of inflammatory cells is verydifferent from the ability to directly chemically affect an oxygenradical. Both will lower inflammation, but only inflammatory regulatorscan up or down regulate the level of inflammation.

Pyruvates and pyruvate precursors control the positive and negativeeffects of inflammatory agents such as cytokines, transforming growthfactor-β, elastase, and white blood cells at higher levels. Too high anumber of white blood cells and other inflammatory agents aredetrimental to lungs. Pyruvates and pyruvate precursors will lower andprotect cells and organs from excess inflammatory agents and white bloodcell numbers when they are high and infections are not involved.Moderate to severe asthmatics, emphysema patients, produce much higherlevels of inflammatory agents including oxygen radicals especially insmokers and low dosages of pyruvates produce better results in thesepatients by lowering excess levels of inflammatory agents. The abilityto control the levels of inflammation is important. Over production orunder production is detrimental and produces various diseases in boththe lungs and nasal cavities. Dosages of 5 ml of 0.5 mM pyruvates reducethe inflammatory markers in patients with lung diseases and can be usedin diseases where inflammation is a problem, i.e., in smokers (21), mildasthmatics (21), in intubated or tracheostomized patients (19), innormal subjects after exercise and hyperventilation (21), COPD patients(22), and in patients with cystic fibrosis (22) with kartagener'ssyndrome (22), moderate or severe asthma (22), Sarcoidosis (22), andfibrosing alveolitis (22). Increased levels of inflammatory cytokinesespecially IL-8 which is a neutrophil activating cytokine, arechemotactic for eosinophils, which produce and enhance inflammation(20). Acute treatment with corticosteroids during an exacerbation ofasthma is associated with a decline in inflammatory markers in adultsand children (23).

In contrast, higher dosages of pyruvates can increase the number ofwhite blood cells and the synthesis of cytokines needed in diseaseswhere cytokines are abnormally low, such as in infections, cancer, andwith the use of drugs. In all lung diseases, when inflammation is high,some cytokines are suppressed. Dosages of 5 ml of 5 mM pyruvates orhigher beyond that needed to neutralize oxygen radicals will enter thebronchial and lung cells and increase the levels of white blood cellsand IL-1, IL-6, IL-8, IL-10, TNF-α, and elastase to help fightinfections and bring balance to the immune system. All cells have atransport system that allows cells to concentrate pyruvate at higherconcentrations than serum levels. In the cell, pyruvate raises the pHlevel, increases levels of ATP, decreasing levels of ADP and cAMP, andincreases levels of GTP, while decreasing levels of cGMP.

In summary, pyruvate enhances the body's ability to increaseinflammatory agents to fight infections and tumors or to bring balanceto the pulmonary or sinus immune system. During inflammation certaincytokines are elevated, whereas certain other cytokines are suppressed.The use of high levels of pyruvate alone or in combination with otherdrugs are effective for the treatment of lung diseases such as asthma,COPD and, in the treatment of tumors, bacterial infections, fungalinfections, viral infections, angina, ischemic diseases, allergicrhinitis, sinusitis and congestive heart failure, where inflammatoryagents and certain cytokines are low or suppressed.

The pulmonary diseases suitable for treatment by the cytokine regulatorsof the present invention include, but are not limited to, bronchialasthma, acute bronchitis, emphysema, chronic obstructive emphysema,chronic obstructive pulmonary disease, centrilobular emphysema,panacinar emphysema, chronic obstructive bronchitis, smokers disease,reactive airway disease, cystic fibrosis, bronchiectasis, acquiredbronchiectasis, kartaagener's syndrome, acelectasis, acute atelectasis,chronic acelectasis, pneumonia, essential thrombocytemia, legionnaire'sdisease, psittacosis, fibrogenic dust disease, hypersensitivity diseasesof the lung, idiopathic infiltrative diseases of the lungs, chronicobstructive pulmonary disorder, adult respiratory distress syndrome,pulmonary tumors, and diseases caused by organic dust, irritant gases,(Allergic Rhinitis, Sinusitis) and chemicals. Preferred disease statesare cystic fibrosis, bronchial asthma, and chronic obstructive pulmonarydisease, Allergic Rhinitis, and sinusitis.

The pulmonary tumors suitable for treatment by the cytokine regulatorsof the present invention include, but are not limited to, epidermoid(squamous cell) carcinoma, small cell (oat cell) carcinoma,adenocarcinoma, and large cell (anaplastic) carcinoma.

The inflammatory agent in the present invention may be selected from awide variety of inflammatory agents. Preferred inflammatory agents arecytokines, transforming growth factor-β, elastase, and white bloodcells. Preferred cytokines may be selected from the group consisting ofinterleukin-1, interleukin-2, interleukin-4, interleukin-6,interleukin-8, interleukin-10, interleukin-17, and interleukin-23. Morepreferred cytokines are interleukin-1, interleukin-6, interleukin-8 andinterleukin 10. IL-10, IL-17, and IL-23 are all regulated by the levelsof IL-6 and IL-8 and so regulation of IL-6 and IL-8 can regulate IL-10,IL-17, and IL-23.

Another preferred cytokine is tumor necrosis factor-α. Tumor necrosisfactor-α is a cytokine involved in systemic inflammation and is a memberof a group of cytokines that all stimulate the acute phase reaction.

Another preferred cytokine is interferon-α and interferon-β. Interferonsare glycoproteins that assist the immune response by inhibiting viralreplication within other cells of the body.

Another preferred inflammatory agent is transforming growthfactor-β(TGF-β). Transforming growth factor-β regulates growth andproliferation of cells, blocking the growth of many different cell typesincluding tumor cells.

Another preferred inflammatory agent is elastase. Elastase is an enzymethat digests and degrades a number of proteins including elastin, anelastic substance found in the lungs and other organs.

Another preferred inflammatory agent is white blood cells. White bloodcells or leukocytes are cells of the immune system, which defend thebody against both infectious disease and foreign materials. Severaldifferent and diverse types of leukocytes exist, however they are allproduced and derived from a pluripotent cell in the bone marrow known asa hematopoietic stem cell. Leukocytes are found throughout the body,including the blood and lymphatic system.

The inflammatory regulators in the present invention are pyruvates andpyruvate precursors. Non-limiting illustrative examples of pyruvatesinclude pyruvic acid, lithium pyruvate, sodium pyruvate, potassiumpyruvate, magnesium pyruvate, calcium pyruvate, zinc pyruvate, manganesepyruvate, aluminum pyruvate, ammonium pyruvate, and mixtures thereof.Non-limiting illustrative examples of pyruvate precursors includepyruvyl-glycine, pyruvyl-alanine, pyruvyl-leucine, pyruvyl-valine,pyruvyl-isoleucine, pyruvyl-phenylalanine, pyruvamide, salts of pyruvicacid, and mixtures thereof.

The amount of the inflammatory regulator present in the therapeuticcompositions of the present invention is a therapeutically effectiveamount. A therapeutically effective amount of the inflammatory regulatoris that amount of the inflammatory agent necessary to treat thepulmonary disease. The exact amount of inflammatory regulator is amatter of preference subject to such factors as the type of inflammatoryregulator being employed, the type of condition being treated as well asthe other ingredients in the composition. The exact amount ofinflammatory regulator will also be determined by whether the pulmonarydisease is infected or uninfected. In general, the dosage of theinflammatory regulator may range from about 0.0001 mg to about 1 gram,preferably from about 0.001 mg to about 0.8 gram, and more preferablyfrom about 0.01 mg to about 0.6 gram.

In another embodiment, the pyruvate or pyruvate precursor inflammatoryregulator further may further comprise α-keto-isovaleric acid, or aprecursor thereof. In general, the dosage of α-keto-isovaleric acid mayrange from about 0.0001 mg to about 1 gram, preferably from about 0.001mg to about 0.8 gram, and more preferably from about 0.01 mg to about0.6 gram.

In one embodiment, the level of inflammatory agents in the mammaliancells is abnormally low in the disease state. In another embodiment, thelevel of inflammatory agents in the mammalian cells is abnormally highin the disease state. Whether the levels of inflammatory agents that areabnormally low or abnormally high can be determined from the level ofinflammatory agents in a patient's lungs and sputum.

In many cases, pulmonary diseases produce infections that theseinflammatory regulators can treat. Such infections may be bacterial,viral, or fungal. The inflammatory regulators may be inhaled first toregulate inflammatory agents followed by inhalation of a therapeuticagent. The therapeutic agent may be administered prior to, concomitantlywith, or after administration of the inflammatory regulator. Thetherapeutic agent may be selected from the group consisting ofantibacterials, antivirals, antifungals, antitumors, antihistamines,proteins, enzymes, hormones, nonsteroidal anti-inflammatories,cytokines, nicotine, insulin, and steroids. All these therapeutic agentselicit an immune response, which will cause a suppression of certaincytokines.

The antibacterial agents which may be employed in the therapeuticcompositions may be selected from a wide variety of water-soluble andwater-insoluble drugs, and their acid addition or metallic salts, usefulfor treating pulmonary diseases. Both organic and inorganic salts may beused provided the antibacterial agent maintains its medicament value.The antibacterial agents may be selected from a wide range oftherapeutic agents and mixtures of therapeutic agents, which may beadministered in sustained release or prolonged action form. Nonlimitingillustrative specific examples of antibacterial agents include bismuthcontaining compounds, sulfonamides; nitrofurans, metronidazole,tinidazole, nimorazole, benzoic acid; aminoglycosides, macrolides,penicillins, polypeptides, tetracyclines, cephalosporins,chloramphenicol, and clindamycin. Preferably, the antibacterial agent isselected from the group consisting of bismuth containing compounds, suchas, without limitation, bismuth aluminate, bismuth subcitrate, bismuthsubgalate, bismuth subsalicylate, and mixtures thereof; thesulfonamides; the nitrofurans, such as nitrofurazone, nitrofurantoin,and furozolidone; and miscellaneous antibacterials such asmetronidazole, tinidazole, nimorazole, and benzoic acid; andantibiotics, including the aminoglycosides, such as gentamycin,neomycin, kanamycin, and streptomycin; the macrolides, such aserythromycin, clindamycin, and rifamycin; the penicillins, such aspenicillin G, penicillin V, Ampicillin and amoxicillin; thepolypeptides, such as bacitracin and polymyxin; the tetracyclines, suchas tetracycline, chlorotetracycline, oxytetracycline, and doxycycline;the cephalosporins, such as cephalexin and cephalothin; andmiscellaneous antibiotics, such as chloramphenicol, and clindamycin.More preferably, the antibacterial agent is selected from the groupconsisting of bismuth aluminate, bismuth subcitrate, bismuth subgalate,bismuth subsalicylate, sulfonamides, nitrofurazone, nitrofurantoin,furozolidone, metronidazole, tinidazole, nimorazole, benzoic acid,gentamycin, neomycin, kanamycin, streptomycin, erythromycin,clindamycin, rifamycin, penicillin G, penicillin V, Ampicillinamoxicillin, bacitracin, polymyxin, tetracycline, chlorotetracycline,oxytetracycline, doxycycline, cephalexin, cephalothin, chloramphenicol,Mupericin and clindamycin.

The amount of antibacterial agent which may be employed in thetherapeutic compositions of the present invention may vary dependingupon the therapeutic dosage recommended or permitted for the particularantibacterial agent. In general, the amount of antibacterial agentpresent is the ordinary dosage required to obtain the desired result.Such dosages are known to the skilled practitioner in the medical artsand are not a part of the present invention. In a preferred embodiment,the antibacterial agent in the therapeutic composition is present in anamount from about 0.01% to about 10%, preferably from about 0.1% toabout 5%, and more preferably from about 1% to about 3%, by weight.

The antiviral agents which may be employed in the therapeuticcompositions may be selected from a wide variety of water-soluble andwater-insoluble drugs, and their acid addition or metallic salts, usefulfor treating pulmonary diseases. Both organic and inorganic salts may beused provided the antiviral agent maintains its medicament value. Theantiviral agents may be selected from a wide range of therapeutic agentsand mixtures of therapeutic agents, which may be administered insustained release or prolonged action form. Nonlimiting illustrativecategories of such antiviral agents include RNA synthesis inhibitors,protein synthesis inhibitors, immunostimulating agents, proteaseinhibitors, and cytokines. Nonlimiting illustrative specific examples ofsuch antiviral agents include the following medicaments.

(a) Acyclovir (9-[(2-hydroxyethyloxy)methyl]guanine, ZOVIRAX®) is awhite, crystalline powder with a molecular weight of 225 Daltons and amaximum solubility in water of 2.5 mg/mL at 37° C. Acyclovir is asynthetic purine nucleoside analogue with in vitro and in vivoinhibitory activity against human herpes viruses including herpessimplex types 1 (HSV-1) and 2 (HSV-2), varicella-zoster virus (VZV),Epstein-Barr virus (EBV), and cytomegalovirus (CMV).(b) Foscarnet sodium (phosphonoformic acid trisodium salt, FOSCAVIR) isa white, crystalline powder containing 6 equivalents of water ofhydration with an empirical formula of Na₃CO₆P₆H₂O and a molecularweight of 300.1. Foscarnet sodium has the potential to chelate divalentmetal ions such as calcium and magnesium, to form stable coordinationcompounds. Foscarnet sodium is an organic analogue of inorganicpyrophosphate that inhibits replication of all known herpes viruses invitro including cytomegalovirus (CMV), herpes simplex virus types 1 and2 (HSV-1, HSV-2), human herpes virus 6 (HHV-6), Epstein-Barr virus(EBV), and varicella-zoster virus (VZV). Foscarnet sodium exerts itsantiviral activity by a selective inhibition at the pyrophosphonatebinding site on virus-specific DNA polymerases and reversetranscriptases at concentrations that do not affect cellular DNApolymerases.(c) Ribavirin (1-β-D-ribofuranosyl-1,2,4-triazole-3-carboxamide,VIRAZOLE®) is a synthetic nucleoside which is a stable, white,crystalline compound with a maximum solubility in water of 142 mg/ml at25° C. and with only a slight solubility in ethanol. The empiricalformula is C₈H₁₂N₄O₆ and the molecular weight is 244.2 Daltons.Ribavirin has antiviral inhibitory activity in vitro against respiratorysyncytial virus, influenza virus, and herpes simplex virus. Ribavirin isalso active against respiratory syncytial virus (RSV) in experimentallyinfected cotton rats. In cell cultures, the inhibitory activity ofRibavirin for RSV is selective. The mechanism of action is unknown.Reversal of the in vitro antiviral activity by guanosine or xanthosinesuggests Ribavirin may act as an analogue of these cellular metabolites.(d) Vidarabine (adenine arabinoside, Ara-A,9-β-D-arabinofuranosyladenine monohydrate, VIRA-A®) is a purinenucleoside obtained from fermentation cultures of Streptomycesantibiotics. Vidarabine is a white, crystalline solid with the empiricalformula, C₁₀H₁₃N₆O₄H₂O. The molecular weight of vidarabine is 285.2, thesolubility is 0.45 mg/ml at 25° C., and the melting point ranges from260° C. to 270° C. Vidarabine possesses in vitro and in vivo antiviralactivity against Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2),and in vitro activity against varicella-zoster virus (VZV). Theantiviral mechanism of action has not yet been established. Vidarabineis converted into nucleotides, which inhibit viral DNA polymerase.(e) Ganciclovir sodium (9-(1,3-dihydroxy-2-propoxymethyl)guanine,monosodium salt, CYTOVENE®, CYMEVENE®) is an antiviral drug activeagainst cytomegalovirus for intravenous administration. Ganciclovirsodium has a molecular formula of C₆H₁₂N₆NaO₄ and a molecular weight of277.21. Ganciclovir sodium is a white lyophilized powder with an aqueoussolubility of greater than 50 mg/mL at 25° C. Ganciclovir is a syntheticnucleoside analogue of 2′-deoxyguanosine that inhibits replication ofherpes viruses both in vitro and in vivo. Sensitive human virusesinclude cytomegalovirus (CMV), herpes simplex virus-1 and -2 (HSV-1,HSV-2), Epstein-Barr virus (EBV), and varicella zoster virus (VZV).(f) Zidovudine [azidothymidine (AZT), 3′-azido-3′-deoxythymidine,RETROVIR®] is an antiretroviral drug active against humanimmunodeficiency virus (HIV) for oral administration. Zidovudine is awhite to beige, odorless, crystalline solid with a molecular weight of267.24 Daltons and a molecular formula of C₁₀H₁₃N₅O₄. Zidovudine is aninhibitor of the in vitro replication of some retroviruses including HIV(also known as HTLV III, LAV, or ARV). Zidovudine is a thymidineanalogue in which the 3′-hydroxy (—OH) group is replaced by an azido(—N₃) group.(g) Phenol (carbolic acid) is a topical antiviral, anesthetic,antiseptic, and antipruritic drug. Phenol is a colorless or whitecrystalline mass, which is soluble in water, has a characteristic odor,a molecular formula of C₆H₆O, and a molecular weight of 94.11.(h) Amantadine hydrochloride (1-adamantanamine hydrochloride,SYMMETREL®) has pharmacological actions as both an anti-Parkinson and anantiviral drug. Amantadine hydrochloride is a stable white or nearly,white crystalline powder, freely soluble in water and soluble in alcoholand in chloroform. The antiviral activity of amantadine hydrochlorideagainst influenza A is not completely understood but the mode of actionappears to be the prevention of the release of infectious viral nucleicacid into the host cell.(i) Interferon α-n3 (human leukocyte derived, ALFERON®) is a sterileaqueous formulation of purified, natural, human interferon α proteinsfor use by injection. Interferon α-n3 injection consists of interferon αproteins comprising approximately 166 amino acids ranging in molecularweights from 16,000 to 27,000 Daltons. Interferons are naturallyoccurring proteins with both antiviral and antiproliferative properties.(j) Interferon α-2a (recombinant, ROFERON-A®) is a sterile proteinproduct for use by injection. Interferon α-2a is a highly purifiedprotein containing 165 amino acids, and it has an approximate molecularweight of 19,000 Daltons. The mechanism by which Interferon α-2a,recombinant, exerts antitumor or antiviral activity is not clearlyunderstood. However, it is believed that direct antiproliferative actionagainst tumor cells, inhibition of virus replication, and modulation ofthe host immune response play important roles in antitumor and antiviralactivity.(k) Oseltamivir((3R,4R,5S)-4-acetylamino-5-amino-3-(1-ethylpropoxy)-1-cyclohexene-1-carboxylicacid ethyl ester, TAMIFLU®) is a is an antiviral drug that is used inthe treatment and prophylaxis of both influenza virus A and Influenzavirus B. Oseltamivir is a neuraminidase inhibitor. It acts as atransition-state analogue inhibitor of influenza neuraminidase,preventing new viruses from emerging from infected cells. Oseltamivirhas a molecular formula of C₁₆H₂₈N₂O₄.

Preferred antiviral agents to be employed may be selected from the groupconsisting of acyclovir, foscarnet sodium, Ribavirin, vidarabine,Ganciclovir sodium, zidovudine, phenol, amantadine hydrochloride, andinterferon α-n3, interferon α-2a, and Oseltamivir. In a preferredembodiment, the antiviral agent is selected from the group consisting ofacyclovir, foscarnet sodium, Ribavirin, vidarabine, and Ganciclovirsodium. In a more preferred embodiment, the antiviral agent isacyclovir.

The amount of antiviral agent which may be employed in the therapeuticcompositions of the present invention may vary depending upon thetherapeutic dosage recommended or permitted for the particular antiviralagent. In general, the amount of antiviral agent present is the ordinarydosage required to obtain the desired result. Such dosages are known tothe skilled practitioner in the medical arts and are not a part of thepresent invention. In a preferred embodiment, the antiviral agent in thetherapeutic composition is present in an amount from about 0.1% to about20%, preferably from about 1% to about 10%, and more preferably fromabout 2% to about 7%, by weight.

The antifungal agents which may be employed in the therapeuticcompositions may be selected from a wide variety of water-soluble andwater-insoluble drugs, and their acid addition or metallic salts, usefulfor treating pulmonary diseases. Both organic and inorganic salts may beused provided the antifungal agent maintains its medicament value. Theantifungal agents may be selected from a wide range of therapeuticagents and mixtures of therapeutic agents, which may be administered insustained release or prolonged action form. Nonlimiting illustrativespecific examples of antifungal agents include the followingmedicaments: miconazole, clotrimazole, tioconazole, terconazole,povidone-iodine, and butoconazole. Other antifungal agents are lacticacid and sorbic acid. Preferred antifungal agents are miconazole andclotrimazole.

The amount of antifungal agent, which may be employed in the therapeuticcompositions of the present invention may vary depending upon thetherapeutic dosage recommended or permitted for the particularantifungal agent. In general, the amount of antifungal agent present isthe ordinary dosage required to obtain the desired result. Such dosagesare known to the skilled practitioner in the medical arts and are not apart of the present invention. In a preferred embodiment, the antifungalagent in the therapeutic composition is present in an amount from about0.05% to about 10%, preferably from about 0.1% to about 5%, and morepreferably from about 0.2% to about 4%, by weight.

The antitumor agents which may be employed in the therapeuticcompositions may be selected from a wide variety of water-soluble andwater-insoluble drugs, and their acid addition or metallic salts, usefulfor treating pulmonary diseases. Both organic and inorganic salts may beused provided the antitumor agent maintains its medicament value. Theantitumor agents may be selected from a wide range of therapeutic agentsand mixtures of therapeutic agents, which may be administered insustained release or prolonged action form. Nonlimiting illustrativespecific examples include anti-metabolites, antibiotics, plant products,hormones, and other miscellaneous chemotherapeutic agents. Chemicallyreactive drugs having nonspecific action include alkylating agents andN-alkyl-N-nitroso compounds. Examples of alkylating agents includenitrogen mustards, azridines (ethylenimines), sulfonic acid esters, andepoxides.

Anti-metabolites are compounds that interfere with the formation orutilization of a normal cellular metabolite and include amino acidantagonists, vitamin and coenzyme antagonists, and antagonists ofmetabolites involved in nucleic acid synthesis such as glutamineantagonists, folic acid antagonists, pyrimidine antagonists, and purineantagonists. Antibiotics are compounds produced by microorganisms thathave the ability to inhibit the growth of other organisms and includeactinomycins and related antibiotics, glutarimide antibiotics,sarkomycin, fumagillin, streptonigrin, tenuazonic acid, actinogan,peptinogan, and anthracyclic antibiotics such as doxorubicin. Plantproducts include colchicine, podophyllotoxin, and vinca alkaloids.Hormones include those steroids used in breast and prostate cancer andcorticosteroids used in leukemias and lymphomas. Other miscellaneouschemotherapeutic agents include urethane, hydroxyurea, and relatedcompounds; thiosemicarbazones and related compounds; phthalanilide andrelated compounds; and triazenes and hydrazines. The anticancer agentmay also be a monoclonal antibody or the use of X-rays. In a preferredembodiment, the anticancer agent is an antibiotic. In a more preferredembodiment, the anticancer agent is doxorubicin. In a most preferredembodiment, the anticancer agent is doxorubicin.

The amount of antitumor agent, which may be employed in the therapeuticcompositions of the present invention may vary depending upon thetherapeutic dosage recommended or permitted for the particular antitumoragent. In general, the amount of antitumor agent present is the ordinarydosage required to obtain the desired result. Such dosages are known tothe skilled practitioner in the medical arts and are not a part of thepresent invention. In a preferred embodiment, the antitumor agent in thetherapeutic composition is present in an amount from about 1% to about50%, preferably from about 10% to about 30%, and more preferably fromabout 20% to about 25%, by weight.

The therapeutic agent of the present invention may also be a nasallyadministered steroid which when administered with pyruvate and/or one ofthe other α-keto acids provides immediate and long term systemic reliefof allergic rhinitis, sinusitis and diseases caused by organic dust,irritant gases, and chemicals. Steroids useful in the practice of thepresent invention include those selected from the group consisting offluticasone, (Flonase®), budesonide (Rhinocort®), beclomethasone,mometasone, flunisolide, triamcinolone and mixtures thereof. These mayalso be used in combination with an antihistamine. Suitableantihistamines are selected from the group consisting ofpseudoephedrine, loratadine, fexofenadine, diphenhydramine, famodidine,ranitidine, citirazine, and other H₁ and H₂ antagonists. Theanti-histamines may actually be administered singly and alone with thepyruvate compositions of the present invention in or together with thesteroid for enhanced nasal relief.

Nicotine is an alkaloid found predominantly in tobacco and constitutesabout 0.6-3% of tobacco by dry weight. Cigarette smoking which containsnicotine has been shown to suppress the immune system to causeinfections and problems in the lungs and sinuses. In low concentrations,an average cigarette yields about 1.0 mg of absorbed nicotine. Nicotineacts as a stimulant in mammals and is one of the main factorsresponsible for the dependence-forming properties of tobacco smoking.Nicotine is a hygroscopic, oily liquid that is miscible with water inits base form. As a nitrogenous base, nicotine forms salts with acidsthat are usually solid and water-soluble. The primary therapeutic use ofnicotine is in treating nicotine dependence in order to eliminatesmoking with its health risks. In low concentrations, an averagecigarette yields about 1.0 mg of absorbed nicotine. Nicotine acts as astimulant in mammals and is one of the main factors responsible for thedependence-forming properties of tobacco smoking. Nicotine is ahygroscopic, oily liquid that is miscible with water in its base form.The primary therapeutic use of nicotine is in treating nicotinedependence in order to eliminate smoking with its health risks.

Insulin is an animal hormone, produced in the pancreas, whose presenceinforms the body's cells that the animal is well fed, causing liver andmuscle cells to take in glucose and store it in the form of glycogen,and causing fat cells to take in blood lipids and turn them intotriglycerides. Insulin is used medically to treat some forms of diabetesmellitus. Patients with type 1 diabetes mellitus depend on externalinsulin (most commonly injected subcutaneously) for their survivalbecause of the absence of the hormone. Patients with type 2 diabetesmellitus have insulin resistance, relatively low insulin production, orboth; some type 2 diabetics eventually require insulin when othermedications become insufficient in controlling blood glucose levels.

Insulin is a peptide hormone composed of 51 amino acid residues.Insulin's genetic structure varies marginally between species of animal.Bovine insulin differs from human in only three amino acid residues, andporcine insulin in one. Even insulin from some species of fish issimilar enough to human to be effective in humans. The C-peptide ofproinsulin, however, is very divergent from species to species. Allstructures of insulin useful in humans, including synthetic “human”insulin, may be employed in the present invention. Unlike manymedicines, insulin cannot be taken orally. Like nearly all proteinsintroduced into the gastrointestinal tract, insulin is degraded losingall insulin activity. Insulin is usually taken as subcutaneousinjections or may be inhaled. The delivery of insulin by inhalation isnot new. As with all insulin's, a patients' dosage must be adjusted forhis/her particular situation. There are several ways to doseshort-acting (rapid-acting) insulin. The following table outlinesestimates of the recommended starting inhaled insulin dosages (based onbody weight).

Starting Inhaled Insulin Weight Dose Blisters  66 to 87 pounds 1 mg permeal One 1-mg blister  88 to 132 pounds 2 mg per meal Two 1-mg blisters133 to 176 pounds 3 mg per meal One 3-mg blister 177 to 220 pounds 4 mgper meal One 1-mg blister plus one 3-mg blister 221 to 264 pounds 5 mgper meal Two 1-mg blisters plus one 3-mg blister 265 to 308 pounds 6 mgper meal Two 3-mg blistersIt was surprisingly and un-expectedly discovered that inhaling insulinwith pyruvate reduced the irritation produced from insulin and producedbetter results than the use of insulin by itself. Diabetics that inhaleinsulin have difficulty if they have a pulmonary disease like COPD,which effects the dosage received. The addition of pyruvate enhancedinsulin uptake for patients that have diabetes with pulmonary lung orupper respiratory sinus diseases.

Moreover, the inhalation of insulin is not only useful in the treatmentof diabetes, but it can be used to treat Alzheimer's. In one patientwith diabetes and mild Alzheimer's, the inhalation of insulin withpyruvate improved mental functions over the same concentration ofinhaled insulin alone. It was discovered that inhaled pyruvate mayincrease IL-10 in the brain thus increases the efficacy of inhaledinsulin, increasing cognitive functions in Alzheimer's patients. It hasalso been shown that certain immune system components when increased,enhances cognitive functions and decreased the amyloid protein, known todisrupt cognitive functions.

Obviously, numerous modifications and variations of the presentinvention are possible in the light of the above teachings and theinvention is not limited to the examples herein. It is thereforeunderstood that within the scope of the appended claims, the inventionmay be practiced otherwise than as specifically described herein.Throughout this application, various publications have been referenced.The disclosures in these publications are incorporated herein byreference in order to more fully describe the state of the art. Thecompounds of the present invention can be prepared according to theexamples set out below. The examples are presented for purposes ofdemonstrating, but not limiting, the preparation of the compounds andcompositions of this invention.

EXAMPLES Example 1 Inhaled Sodium Pyruvate for the Treatment of CysticFibrosis Double Blind, Placebo-Controlled, Safety Study SputumInflammatory Biomarkers

All of the enrolled and dosed subjects were able to provide sputumsamples for analysis before and after exposure to study drug (sodiumpyruvate for inhalation at 0.5, 1.5, and 5.0 mM levels). The subjectswere given 5 ml samples to inhale. Specimens were of good quality forthe planned assays.

The 0.5 mM levels of sodium pyruvate using 5 ml samples contain 0.28 mgof sodium pyruvate. The 5 mM levels of sodium pyruvate using 5 mlcontain 2.8 mg of sodium pyruvate.

Samples were divided into two main aliquots after processing. The firstaliquot was left untreated to be able to assay for the activity of freeelastase. The second aliquot was treated with protease inhibitorspheylmethanesulfonylfluoride (PMSF) and ethylenediamine tetraacetic acid(EDTA) to stop any degradation of the cytokines of interest (IL-6, IL-8,IL-10, IL-17, and IL-23) and total protein.

For IL-10, IL-17 and IL-23, levels on sputum were at or below the limitof detection for the assays (7, 2 and 20 pg/mL respectively). For theother markers (total protein, elastase, IL-6, IL-8, TNF-α) levelsdetected where within those typically found in cystic fibrosis (CF)patients. Overall, significant changes were noted in these biomarkerstested in sputum by drug dose level or for the group as a whole (FIGS.1-10).

As set forth above, FIG. 1 is a graph illustrating individual sputumtotal protein levels before and after study drug inhalation. Slash marksrepresent the median level. FIG. 2 is a graph illustrating median changefrom pre- to post-study drug inhalation in sputum total protein levelsand by drug dose level while FIG. 3 is a graph illustrating individualsputum free elastase levels before and after study drug inhalation. FIG.4 is a graph illustrating median change from pre- to post-study druginhalation in sputum free elastase levels and by drug dose level. FIG. 5is a graph illustrating individual sputum IL-6 levels before and afterstudy drug inhalation. FIG. 6 is a graph illustrating median change frompre- to post-study drug inhalation in sputum IL-6 levels and by drugdose level. FIG. 7 is a graph illustrating individual sputum IL-8 levelsbefore and after study drug inhalation. Slash marks represent the medianlevel. FIG. 8 is a graph illustrating median change from pre- topost-study drug inhalation in sputum IL-8 levels and by drug dose level.FIG. 9 is a graph illustrating individual sputum TNF-α levels before andafter study drug inhalation. FIG. 10 is a graph illustrating medianchange from pre- to post-study drug inhalation in sputum TNF-α levelsand by drug dose level.

Given the changes noted in the cell counts (both in peripheral blood andsputum) this finding is intriguing. Since evidence for cellular influxwas noted, it would have been expected to see this paralleled by acorresponding increase in cytokines and particularly in the freeelastase activity. The study drug blocked this pro-inflammatory effect.

The data clearly showed that pyruvate can up or down regulateinflammation depending on concentration. White blood cell counts werereduced 25% with the inhalation of 5 ml of 0.5 mM sodium pyruvate, aswas total proteins, elastase, as were IL-6, IL-8, and TNF-α. White bloodcell counts were increased by 25% with the inhalation of 5 ml of 5 mMpyruvate or higher as was the total proteins, elastase, IL-6, IL-8, andTNF-α.

Tissue Culture Studies

To investigate the ability of pyruvate to regulate the inflammatoryprocess during an infection, the MatTek EpiDerm Assay was used. TheMatTek Epiderm tissue samples were treated with pyruvate and thecombination of pyruvate and α-ketoisovalerate both at 20 mMconcentrations or higher to determine if the combination would regulateIL-I and IL-8 up or down during a simulated infection. Following aone-hour equilibration, the Epiderm tissues were placed into theincubator (37° C., 5% CO₂) in assay medium. The old medium was replacedwith fresh medium and the test articles were applied to the tissuesamples. The test articles remained in contact with the tissue forvarious dosing times, one hour, then at four hours, and at 20 hours. Thetesting was run in duplicate. Various immunostimulators sodium dodecylsulfate (SDS), glycoprotein D (gpD) were used singly or with the α-ketoacids to replicate an infection, along with vehicle controls. Untreatedsamples were used as negative controls. Following treatment, the mediafrom the tissues samples were tested in Elisa kits for IL-1 and IL-8according to the manufacture's protocols.

The 0.5 mM levels of sodium pyruvate using 5 ml samples contain 0.28 mgof sodium pyruvate. The 10 mM levels of sodium pyruvate using 5 mlcontain 5.6 mg of sodium pyruvate. The 20 mM levels of sodium pyruvateusing 5 ml contain 11.2 mg of sodium pyruvate. The 40 mM levels ofsodium pyruvate using 5 ml contain 22.4 mg of sodium pyruvate.

A quantity of 5 ml of 0.1 mM to 100 mM of α-keto isovalerate was used. Aquantity of 5 ml of 20 mM of α-keto isovalerate contains 13.8 mg. Aquantity of 5 ml of 40 mM of α-keto isovalerate contains 27.6 mg. Aquantity of 5 ml of 100 mM of α-keto isovalerate contains 69 mg.

Results

The primary end points were the levels of IL-8 and IL-1 after treatmentwith an immunostimulator, pyruvate and the combination of pyruvate andα-ketoisovalerate. The immunostimulator did not increase the cytokinesby themselves. This model did not have white blood cells to respond tothe immunostimulator or produce oxygen radicals. The α-keto acids didnot increase the cytokines also in this model. The immunostimulators incombination with pyruvate and α-ketoisovalerate increased IL-8 over300%, which shows direct antimicrobial activity, compared to theuntreated controls. IL-8 activates neutrophils to increase their numbersat the infected site. In the same experiment, IL-1 was decreasedsignificantly (over 200%). IL-1 increases inflammation and decreaseshealing times. This test clearly showed that the α-keto acids regulatedthe inflammatory process in dermal tissues in a manner that wouldincrease the bodies ability to fight infected wounds and increase thebody's ability to healing quicker. The same experiment was done withvirally infected cells and the pyruvate and combination of pyruvate andα-ketoisovalerate decreased viral plaque formation by 50%. Viral plaquesare a direct measure of viral numbers in infected cells. The antiviraldrug, Acyclovir also decreased viral plaques by 60% and the α-keto acidsin combination with acyclovir, totally eliminated the virus from theinfected cells.

Example 2 Inhaled Sodium Pyruvate for the Treatment of Cystic Fibrosisand Other Lung Diseases

Cystic fibrosis (CF) is the most common, lethal inherited disease ofCaucasians. Approximately 30,000 people in the United States and 70,000worldwide have a diagnosis of CF. It is caused by mutations in thecystic fibrosis transmembrane regulator (CFTR) gene. The clinicalmanifestations characteristic of CF include progressive bronchiectaticlung disease with thick mucus production and colonization by Pseudomonasaeruginosa. The CFTR gene mutation results in altered cell transportproperties, which affect both chloride and glutathione secretion.Chronic inflammation, associated with activated neutrophils andmacrophages, is a common feature of CF. Highly reactive toxic oxygen(superoxide anion, free hydroxyl radical, hydrogen peroxide) andnitrogen species (NO, peroxynitrites) are abundant in the chronicinflammatory response in CF and appear to playa prominent role in thepathogenesis of this disease as are excess levels of inflammatorycytokines.

A total of fifteen 15 CF and 10 Asthmatic/COPD patients were treatedwith varying doses of sodium pyruvate. The therapeutic dose, 0.5 mM ofsodium pyruvate lowered the inflammatory cytokines (markers) including,IL-6, IL-8 by 200% or more. IL-10 was not affected. The 5.5 mM or higherconcentrations of sodium pyruvate increased only IL-8. The 10 mMsolution of sodium pyruvate produced the best results by increasing theinflammatory cytokines IL-8 and IL-10 by 200% or more which was asurprise. The 0.5 mm can be used in CF patients to lower inflammationand white blood cell numbers and the 5.5 mM or higher can be used toincrease cytokines and white blood cell numbers needed to fightinfections and balance the immune system, especially if the patient isusing inhaled steroids.

Treatment of HSV-I infected cell with various α-Keto Acids α-Keto acidsregulation of inflammatory anti viral cytokines at varying mMconcentrations α-Keto Acids tested Alone for their Ability to ReduceViral Plaques (numbers live viruses) Percentage of viral plaquereduction in virally infected cells pyruvate + pyruvate + α-keto α-ketoα-keto α-keto pyruvate isovalerate butyrate isovalerate butyrate 05 mM5% 0% 0% 10% 0% 10 mM 10% 5% 0% 20% 3% 20 mM 38% 10% 0% 50% 30% 40 mM50% 20% 5% 74% 40% α-Keto Acids in Combination with Acyclovir(therapeutic dose) Percentage of viral plaque reduction in virallyinfected cells (reduction of live viruses) α-keto Pyruvate isovalerateα-keto butyrate therapeutic dose Acyclovir Acyclovir Acyclovir Acycloviralone 10 mM 66% 42% 39% 40% 20 mM 90% 55% 40% α-Keto Acids Combinationswith Acyclovir Percentage of Viral Plaque Reduction in Virally InfectedCells Pyruvate pyruvate α-keto isovalerate α-keto butyrate AcyclovirAcyclovir 10 mM 78% 63% 20 mM 100% 70%The results clearly show that pyruvate was the only α-keto acid thatincreased the inflammatory cytokines high enough needed to kill highnumbers of the virus in virally infected cells as measured by reductionin viral plaques. Acyclovir a known anti-viral agent, also reduced viralplaques. Unexpectedly, the combination of pyruvate and α-ketoisovalerate produced the best results, by totally eliminating the virusfrom the infected cells. Inhalation of pyruvate at 0.5 mM reduced thelevels of white blood cells, elastase, IL-8, IL-6 and did not increaselevels of TNF-α. Inhalation of more than 5 mM of pyruvate in humansincreased levels of white blood cells elastase, IL-8, IL-6, IL-10 andTNF-α (cytokines) that would be used to kill viruses in the lungs. Thistissue culture data along with data from humans, confirms that someα-keto acids worked and some like α-keto butyrate did not. It appearsthat α-keto butyrate will reduce inflammation and the production ofcytokines, even during an infection.

Example 3 Nicotine/Pyruvate Inhalation Delivery to the Sinuses or Lungs

Nicotine was formulated as a modified sodium pyruvate 20 mM nasalsolution which was then administered as an inhaled mist to the lungs ornasal cavity. A number of clinical test patients squirted each nostrilthree times each, 2 times per day which amounted to 2.94 mg-4.2 mg ofpyruvate and approximately 0.020 mg to 0.5 mg of nicotine per dailydose. A patient that used Nicotrol (nicotine nasal spray) for smokingcessation was treated with a nasal spray of sodium pyruvate 20 mM to 40mM prior to using his nasal inhaler. Each metered dose of Nicotroldelivers 0.5 mg of nicotine. The administration of the 20 mM spray ofnicotine/sodium pyruvate solution allowed the patient to control hisnicotine addiction using less medication and eliminated the irritationand nose soreness associated with nasal nicotine formulations such asNicotrol. Nicotine inhalation can increase the risks of nasal and lunginfections and nicotine can suppress certain components of the immunesystem. The 20 mM pyruvate solution in combination with nicotine reducedthe negative effect of nicotine and enhanced the effect of nicotine as astimulant. High levels of pyruvate, 5.5 mM to 40 mM or higherup-regulate the immune system and its inflammatory agents to balance outthe negative suppressive effects of inhaled nicotine. It was found thatthe inhaled nicotine/pyruvate solution, delivered nasal or directly tothe lungs, was well tolerated over nicotine by itself, which wasirritating. The combination also proved to be synergistic in theobserved effects. On a scale of 1-10 with 10 being very irritating and 1being non-irritating, nicotine solutions delivered without pyruvate wererated a score of 8.0 by 6 patients and those nicotine solutions withpyruvate were rated a 2.0 by the same patients, a 60% reduction inirritation. Smokers who used this formula rated the pyruvate/nicotineformula more efficacious in the ability to control their smoking andsince it was much less irritating and “patient-friendly”, it was ratedas being much more effective than nicotine alone.

Example 4 Insulin/Pyruvate Inhalation in the Sinuses or Lungs

Insulin is an animal hormone, produced in the pancreas, whose presenceindicates to the body and the body's cells that the animal is well fed,causing liver and muscle cells to take in glucose and store it in theform of glycogen, and fat cells to take in blood lipids and turn theminto triglycerides. Insulin is used to treat some forms of diabetesmellitus. Patients with type-1 diabetes mellitus depend on externalinsulin (most commonly injected subcutaneously) for their survivalbecause of the absence of the hormone. Patients with type-2 diabetesmellitus have insulin resistance, relatively low insulin production, orboth; some type 2 diabetics eventually require insulin when othermedications become insufficient in controlling blood glucose levels.Chemically, insulin is a peptide hormone comprised of 51 amino acidresidues and its' structure varies marginally between species of animal.Bovine insulin differs from human in only three amino acid residues, andporcine insulin in one. Even insulin from some species of fish issimilar enough to human to be effective in humans. The C-peptide ofproinsulin, however, is very divergent from species to species. Allstructures of insulin useful in humans, including synthetic “human”insulin, may be employed in the present invention however, insulincannot be taken orally. When prescribed, every individual's dose must beadjusted for that individual's particular situation. There are severalways to dose short-acting as well as rapid-acting insulin. The followingtable outlines estimates of the recommended starting inhaled insulindosages (based on body weight).

Starting Inhaled Insulin Weight Dose Blisters  66 to 87 pounds 1 mg permeal One 1-mg blister  88 to 132 pounds 2 mg per meal Two 1-mg blisters133 to 176 pounds 3 mg per meal One 3-mg blister 177 to 220 pounds 4 mgper meal One 1-mg blister plus one 3-mg blister 221 to 264 pounds 5 mgper meal Two 1-mg blisters plus one 3-mg blister 265 to 308 pounds 6 mgper meal Two 3-mg blistersIt was surprisingly discovered that inhaling insulin with pyruvatereduced the irritation produced from insulin and produced better resultsthan the use of insulin by itself. Diabetics that inhale insulin havedifficulty if they have a pulmonary disease like COPD, which effects thedosage received. The addition of pyruvate enhanced insulin uptake forpatients that have diabetes with pulmonary lung or sinus diseases. Theinhalation of insulin cannot only be used to treat diabetics, but can beused to treat Alzheimer's disease as well. In one patient with diabetesand mild Alzheimer's the inhalation of insulin with pyruvate improvedmental functions over the same concentration of inhaled insulin alone.It was discovered that inhaled pyruvate may increase IL-10 production inthe brain thus increasing the efficacy of inhaled insulin, therebyincreasing the cognitive functions in Alzheimer's patients. It has alsobeen shown that certain immune system components when increased,enhances cognitive functions and decreased the amyloid protein, known todisrupt cognitive functions.

Preliminary Cognitive Testing: Three-Word Recall, “Mini-Cog” and CoinCounting

The following two tests can help gauge memory function when patientsexpress concern. The tests also may help identify patients who need morethorough evaluation. All of these tests are relatively free of influenceby educational level:

1. A Three-Word Delayed Recall Exercise

-   -   The patient is told to remember three words, these being three        common nouns, such as horse, pencil and rose. The patient was        then asked to repeat them.    -   About five minutes later, the patient was then asked recall        them.    -   Individuals without impairment should be able to remember all        three words, especially with such prompts as, “The first word        was the name of an animal,”    -   Remembering only one or two words indicates a need for further        evaluation.        2. The “Mini-Cog” Test, Combining Three-Word Recall with        Clock-Drawing    -   Three simple nouns were given and the patient was asked to        repeat them,    -   The patient was then asked to draw the face of a clock on a        sheet of paper, showing the time as 10 minutes past 11.    -   After the clock has been drawn, ask the patient to repeat the        three words.        -   Patients who remember all three words have no dementia.        -   Patients who remember none of the words should receive            further evaluation,        -   if the patient remembers one or two words, the physician            should refer to the score on the clock drawing to help            interpret this            -   normal clock=non-demented            -   abnormal clock=further evaluation needed        -   Patients who recall all three words but have a problem with            the clock may also require further evaluation.    -   More about clock-drawing and scoring        -   When stating the time to be shown on the clock, avoid            referring to the “hands” of the clock to avoid prompting.            Rather, say “Show the time as 10 past 11.”            -   “10 past 11” tests the ability to translate “10 past”                into the right numerical value.            -   It also requires the use of both halves of the clock                face.        -   There are several scoring systems. A simple one is based on            four points, with a lower score suggesting further            evaluation.            -   One point is given for drawing a closed circle. Some                clinicians prefer to give patients a pre-drawn circle,                so that any accidental distortions in shape do not                affect the placement of the numbers.            -   One point is awarded for including the 12 correct                numbers.            -   One point is given for putting the numbers in the                correct position.            -   One point is awarded for drawing the hands to show the                correct time.

Using the above described tests, on this diabetic Alzheimer's patient,the inhalation of insulin by itself improved cognitive functionmoderately, over base scores, with the patient being able to increasehis scores by 40% over the non use of insulin. When pyruvate wasadministered by itself his scores increased by 45% over basemeasurements. The combination of insulin and pyruvate increased hiscognitive abilities to nearly 90%, showing that pyruvate and insulin aresynergistic.

In the present example, insulin was administered to the lungs of eachpatient in a 5 mL 20 mM pyruvate solution. For the nasal cavity, eachnostril of the patient was squirted three times each, 2 times per daywhich comprises 2.94 mg of pyruvate. Insulin inhalation can increase therisks of nasal and lung infections and insulin can suppress certaincomponents of the immune system. The 20 mM pyruvate solution incombination with insulin balanced the negative effect of insulin andenhanced the effect of insulin. High levels of pyruvate, 5.5 mM to 40 mMor higher up-regulate certain components of the immune system and itsinflammatory agents to balance out the negative suppressive effects ofinhaled insulin and to enhance its uptake. Many investigators have shownthat inhaled insulin can suppress the immune system, thus the use ofhigh levels of pyruvate was shown to up-regulate key components of theimmune system and reduce the inhibitory effects of insulin on theproduction of key inflammatory agents. In subsequent experiments, highlevels of inhaled pyruvate over 5 mM (10-20 mM) has been shown toincrease IL-10 in both the sinuses and lungs.

Example 5

Thirty-nine (39) subjects who were regular nasal spray users due tochronic sinusitis or allergic rhinitis and other sinus diseases wererecruited for a one week open label, in home-use trial. Allergicrhinitis and other sinus diseases result in the elevation of certaininflammatory components and the concurrent down regulation of some otherinflammatory components. When this happens, infections can occur. Theability to up-regulate these immune components helps balance the nasalimmune system to inhibit infections and minimize injury to surroundingtissues. Six (6) different sodium pyruvate+saline combination nasalsprays, based on concentrations of saline and sodium pyruvate, wereadministered to the subjects, and each of the test products were used byat least six subjects. The subjects used the test nasal sprays at home(two to three sprays; two or three times a day) for seven days, in placeof their regular nasal spray.

Prior to, and at the end of the study period, the subjects' nostrilswere examined for mucosal fragility, lesions, erythema, and edema usinga rhinoscope.

Thirty-eight subjects completed the study. The data obtained from therhinoscopic examinations indicated that none of the six (6) sodiumpyruvate+saline nasal spray test products induced nasal fragility,lesions, erythema, or edema. The test formula ranged from 0.5 mMsolutions to 20 mM solutions. A 40 mM sodium pyruvate solution wastested and this was also favorably rated by the patients. The 0.5 mM to5 mM pyruvate solutions were rated non-effective to less effective bythe patients when compared to the 20-40 mM pyruvate solutions. In thiscase we delivered the 20 mM solution nasal solution with sodiumpyruvate. The patient squirted each nostril three times each, 2-3 timesper day is 2.94 mg to 3.8 mg of pyruvate delivered per daily dose. Allthe test sprays were preserved with 0.02% benzalkonium chloride.

81% of all subjects said the 0.45% sodium chloride/sodium pyruvate 20 mM(5 ml contains 11 mg of pyruvate) was “Better Than” or “Comparable” totheir present therapy with regard to less “Stinging.” When questioned bythe Investigator, 35 of 39 (90%) subjects stated that thePyruvate+Saline Nasal Sprays “Opened Nasal Passages,” and“Cleared/Reduced Congestion.” This product received an overall rating of8.0, with all characteristic evaluations rated between 7.6 and 8.7(“Likely to Buy”). A nasal wash delivered 10 mls of the 20 mM pyruvatesolution which is 22 mg of pyruvate being delivered.

This study clearly showed that delivery of high levels of sodiumpyruvate can reduce the toxic effects of inhaled pollutants, nasalsensitivity, soreness, mucus congestion, swelling, and enhanced thepatients' ability to sleep. The formulations were soothing to thenostrils, relieving nasal symptoms, of congestion, irritation, redness,snoring.

Example 6

Eighteen subjects who were regular nasal spray users were givenEmphyClear™, a sodium pyruvate/saline nasal spray to use at home two orthree times a day for seven days, in place of their regular nasal spray.Several of these subjects regularly used saline, or OTC nasal products,including antihistimines and several used steroid-based nasal sprays.These patients suffered from nasal diseases caused by dusts, allergiesand pollutants from irritant gases which cause allergic rhinitis,sinusitis, and other nasal diseases. Prior to, and at the end of thestudy period, the subjects' nostrils were examined for mucosalsensitivity, fragility, lesions, erythema, and edema using a rhinoscope.These pre- and post-study nasal characteristics were rated on a fivepoint scale, zero (“none”) to four (“severe”) scale, and compared.

All 18 subjects completed the study, and none opted to return to theirnormal nasal spray therapy during this period. The data obtained fromthe rhinoscopic examinations indicated that the sodium pyruvate/salinenasal spray did not induce dermal irritation and was effective insignificantly (p=0.006) reducing the erythema in subjects who normallyuse either saline or non-saline nasal sprays including steroids whenpre-test ratings were compared to post-test ratings. Further, subjectiveevaluations from the subjects indicated a positive preference for thesodium pyruvate/saline nasal spray, with 83% of all subjects sayingEmphyClear™ was “Better Than” or “Comparable To” their present therapywith regard to “Soothing;” and a like percentage of all subjects sayingEmphyClear™ was “Better Than” their present therapy for relievingsymptoms.

Ninety-four percent (94%) of all subjects said it was “Better Than”their present therapy with regard to less “Stinging.” When questioned bythe Investigator, 17 of 18 subjects stated that EmphyClear™ “OpenedNasal Passages,” and “Cleared Congestion and reduced snoring,moisturized their nasal passages and enhanced their ability to sleep allnight.” These results were consistent whether the subject normally useda saline or steroid-based nasal spray. This study clearly showed thatdelivery of high levels of sodium pyruvate can reduce the toxic effectsof inhaled pollutants, nasal fragility, congestion do to mucus,swelling, and enhanced their sleeping. Five of these patients sufferedcontinuously form various forms of sinusitis, (viral or bacterialinfections) and while using the pyruvate solutions reported that theirsinusitis decreased or disappeared.

Example 7

A consumer at-home trial comprising a three-month evaluation study wasconducted in which thirty-one (31) subjects that normally used nasalsprays were asked to evaluate the three different sodium pyruvatesolution concentrations over a period of three months. The sodiumpyruvate+saline nasal sprays contained 0.45% saline+either 5 mM, 10 mMor 20 mM sodium pyruvate. The subjects were asked to evaluate the threeproducts in terms of congestive relief, and to indicate any preferencesbetween the three concentrations of sodium pyruvate.

The 0.45% saline+20 mM Sodium Pyruvate nasal spray was the preferredproduct with an overall rating of 90%. 29 out of the 31 subjects (94%)stated that this product “opened their nasal passages and cleared theircongestion and allowed them to sleep longer and sounder.”

In all, 88 patients suffering with allergic rhinitis, sinusitis, othersinus diseases, or were smokers, or had various lung diseases, includingCOPD, ILD, with one patient with an HIV infection, were tested withvarious different Sodium Pyruvate formulae (5.5 mM to 40 mM) and 81patients (92%) stated that the Pyruvate+Saline Nasal Sprays opened theirnasal passages and cleared their congestion for over 12 hours andenhanced sleep Five of these patients suffered continuously from variousforms of sinusitis, (viral or bacterial infections) and while using thepyruvate solutions reported that their sinusitis decreased ordisappeared. In each study the sodium pyruvate+saline nasal sprays wereobjectively and subjectively judged to be “Comparable To” or “BetterThan” Saline or Steroid-based commercial nasal sprays. High levels ofinhaled pyruvate selectively up-regulated certain inflammatory agents tofight infections, and to balance out the negative effects ofinflammation. In normal patients that had no lung or sinus disease orinflammation, high levels of inhaled pyruvate produced no effect beyondsoothing the sinuses and relieving snoring. As discussed infra at page2, when some inflammatory agent levels are elevated, others aresuppressed and abnormally low due to disease or inflammation. Forexample, in allergic rhinitis, IL-10 is suppressed, as are some of theother cytokines.

On the other hand, inhaling high doses of pyruvate enhances and balancesthe immune system. This reduces tissue and cellular damage, and reducesor eliminates infections. IL-10 is capable of inhibiting synthesis ofpro-inflammatory cytokines and also displays a potent ability tosuppress the antigen-induced reactions, thus up-regulating IL-10 hasbeen shown to be effective in treating allergic rhinitis. Testing thelungs or sinuses of several patients for increases in cytokines, asoutlined in example 1, demonstrated that IL-10 increased by 380% withthe inhalation of a 20 mM pyruvate solution. The increase in IL-10levels reduced nasal congestion and fragility, irritation and reducedthe allergic reaction to dust, pollen, mold, and other allergens. Thisalso resulted in a reduction in snoring that was subsequently reportedby most patients.

Example 8

Nine regular Flonase® subjects and eight regular Nasacort® subjects whosuffered chronically from allergic rhinitis and sinusitis from dust,allergens, gases, mold, and pollen, were recruited to evaluatecomparable products containing a reduced level of steroids (50-70%reduction in the drug) in a sodium pyruvate nasal solution. Prior tobeginning the study, the subjects were asked to rate their currentproduct on a 10 point visual analogue scale (VAS) with a value of 0being “terrible” and 10 being “excellent” for the following categories:a) soothing the nostrils; b) relieving sinus symptoms; c) stinging ofthe nostrils, and overall rating of satisfaction. The subjects onaverage rated their current products “good,” with little differencebetween the two products except for a trend toward perceived better“soothing” with Nasacort® than with Flonase®.

At the start of the trial, the subjects were blinded regarding theirtest product, and the test product was used exclusively for 14 days. Thesubjects' nostrils were then objectively evaluated using a nasoscope atdays 0, 7, and 14, and physical exams, including vital signs were alsoadministered at this time. During the 14 day test period, the subjectssubjectively evaluated the test product on a daily basis using a10-point VAS questionnaire. The categories included comparison of thetest products to the subjects' normal therapy in their ability to sooththe nostrils, relieve symptoms, cause/reduce stinging, relievecongestion, and quantify usage, and rate the product on an “overall”basis.

After seven and fourteen days, nasoscope evaluations revealed a trend inreduction of aberrant morphologies for the “Reduced-Strength Flonase®”Test product compared to the nasoscope evaluations obtained on Day 0;and a significant reduction in aberrant morphologies for the“Reduced-Strength Nasacort®” Test product. These objective observationsare consistent with the subjective evaluations where the subjects ratedthe “Reduced-Strength Flonase®” product as “Comparable” or “Better” inall categories, and rated “Reduced-Strength Nasacort®” Test product as“Better” in all categories.

The test products were subjectively judged to be comparable or betterthan the Flonase® or Nasacort® that the subjects typically used. Thesubjects did not rate the test products lower than the Flonase® orNasacort® in any category. The test products were rated as “Better” incomparison to the “Soothing,” “Stinging” and “Relief of Symptoms”characteristics of Flonase®, and, with regard to Nasacort®, the subjectsrated the Test product as “Comparable” across all categories after 14days of use. End-of-Trial subjective comments were also highly favorableto the Test products compared to Flonase® and Nasacort®. Additionally,when asked if they might purchase the product, the subjects' averageresult was 5.4±1.0, indicating that the subjects “Might Purchase,” orwere “Likely to Purchase,” the Test product.

An analysis of the results conclusively showed that the“Reduced-Strength Flonase®” and “Reduced-Strength Nasacort®” Testproduct nasal sprays were found to be as effective as the“full-strength” (i.e. commercial) Flonase® and more effective than thecommercial Nasacort® when the reduced commercial “active ingredients”were delivered to the subjects in a sodium pyruvate solution.

The sodium pyruvate and steroids in the nasal spray were found to actsynergistically. By themselves, steroids will shut down the immunesystem and thus can be toxic, irritating, and habit forming whichthereby increases the percentage of infections in patients. Whenformulated in solution with pyruvate, the steroids were found to actsynergistically which enabled the reduction of inhaled steroid levelsand complemented their reactions in the human body. Inhaledcorticosteroids reduces macrophage inflammatoryprotein-1-α-granulocyte-macrophage colony-stimulating factor, cytokinesand interferon-gamma release from alveolar macrophages in asthma, whichincreases infections. Asthma is characterized by a reduced capacity toproduce IL-10.

The 0.5 mM pyruvate/steroid solution that was used to reduceinflammation and inflammatory markers in the lungs, did not up-regulateinflammatory markers in the nasal cavities or lungs, whereas the 5.5 mM(2.9 mg) to 20 mM sodium piruvate/steroid solution did. This wasunexpected. The 5.5 mM to 20 mM pyruvate solutions in combination withreduced steroids balanced their negative effect and enhanced theirefficacy that allowed for the formulation of an efficacious nasal spraywith a reduction of the steroids by 70%. Steroids suppress the immunesystem to the point that patients that use steroids have a very highrate of infection. High levels of pyruvate, i.e. 5.5 mM to 40 mM orhigher, up-regulate the immune system and its inflammatory agents andthis balances the negative immune suppressive side effects of steroids.During allergic rhinitis or other sinus diseases, certain inflammatorycomponents are elevated, whereas certain other inflammatory componentsare down regulated, and in the process infections can occur. The abilityto up-regulate these immune components help balance the nasal and lungimmune system to inhibit infections and minimize injury to surroundingtissues and reduce steroids by 70%. In Allergic Rhinitis, IL-10 issuppressed, as are some of the other cytokines.

Allergic reactions include four types of reactions, i.e., types I, II,Ill and IV. The type I (immediate-type, anaphylactic) allergic reactionis triggered by the reaction-relating-factor immunoglobulin E(hereinafter abbreviated as an IgE antibody). The reaction steps can bedivided roughly into the following three steps. The first step is asensitization step involving IgE antibody production and binding of theresulting IgE antibodies to mast cells or basophiles. The second stepinvolves degranulation of the mast cells or basophiles and release ofchemical mediators. The third steps involves onset of effects of thereleased chemical mediators on the target organs. Thus, the type Iallergic reaction against foreign antigens leads to onset of symptomsthrough the above reaction steps. Only symptomatic treatments byinhibiting the above second and/or third reaction steps have beencarried out to treat allergic diseases. That is, the treatments arecarried out by inhibiting the release of chemical mediators accompanyingthe degranulation and/or by inhibiting allergic reactions induced by thereleased chemical mediators. These symptomatic treatments have beenknown to be effective not only in systemic administration ofanti-allergic agents but also in their topical administration to thenose, etc. However, the effects of the treatments are limited becausethe treatments do not inhibit IgE antibody production which is the basicfirst step of the type I allergic reaction.

However, because the mechanisms of nasal topical IgE antibody productionare not clear, there is no report on effects of nasally topicallyadministered drugs applicable to nasal topical membrane allergicreaction. As described above, there is no satisfactory anti-allergicpharmaceutical compositions that are effective and safe in nasal topicaladministration until now. In doing nasal lavage studies, up-regulatingcertain cytokines, like IL-10, by using high levels of pyruvate has beenshown to inhibit IgE antibody production.

The data and clinical results clearly show that patients who use asteroid/pyruvate nasal formulation will reduce their chances ofinfections and tissue damage. In a similar experiment described above, acommercial Rhinocort® nasal formula (32 mg of budesonide) was dilutedwith saline to deliver 16 mg of budesonide (50% of the commercialformulation) to the 4 patients that use Rhinocort and that suffered withallergic rhinitis and sinusitis, and other nasal inflammatory diseases.These patients rated budesonide a score of 8.0 on a 1-10 irritationscale with 1 being perceived as non-irritating and 10 being perceived asvery irritating to the sinuses. These patients dosed each nostril 2-3times, four to six times daily to achieve efficacy of their currentproduct, which is 24-36 daily squirts, far exceeding the recommended FDAstandards of 240 mg daily amount. When these patients tested the 50%formulation with the 5.1 mM sodium pyruvate solution, they obtained thesame efficacy, but with half the needed amount of the steroid, but stillused 18-24 squirts daily, which exceeds the FDAs' recommended dailylimits. They rated the product a 5. When these patients tested the 50%steroid formula with the 20 mM sodium pyruvate, they rated the product a2 and all the patients recorded a 20%-30% reduction in usage, 8-12squirts daily usage, clearly showing that the 20 mM pyruvate formulationwas synergistic and un expectedly less irritating to the nasal cavitiesand sinuses and did not exceed the FDA limits of daily steroid usage. A20 mM solution nasal solution of sodium pyruvate squirted into eachnostril three times each 2-3 times per day is 2.94 mg to 3.65 mg ofpyruvate delivered per daily dose.

Ribavirin Inhaled Antiviral and Mupericin Inhaled Antibacterial

This medication is an anti-viral drug used to treat infants and youngchildren who have a severe lung infection caused by a certain virus(respiratory syncytial virus-RSV). Nearly all children become infectedwith this virus before they are 3 years old. Most cases are mild and donot require anti-viral drugs. This medication is used to treat severeRSV infections that need treatment in a hospital. This medication isgiven by continuous inhalation, usually for 12 to 18 hours daily for 3to 7 days or as directed by the doctor. A special machine(small-particle aerosol generator) is used to make a mist, which is theninhaled through the mouth or nose. Chest soreness may occur.Redness/irritation of the nasal cavities and eye or eyelid may alsooccur. When five (5) mls. of a 10 mM solution of pyruvate was used priorto the administration of this medication; the patient reported that theirritation and chest soreness was eliminated. High levels of pyruvate5.5 mM or higher have been shown to up-regulate certain components ofthe immune system to kill virally infected cells. Tissue culture studieswith HSV-1, HSV-2 and various rhinoviruses, have shown that 10-40 mMconcentrations of pyruvate reduced viral plaques by 40-60% by increasingthe synthesis of various cytokines. The combination of variousantivirals with pyruvate totally eliminated the virus infection incells. The inhalation of 5.0 mls of a 20 mM solution of sodium pyruvateused by itself reduced viral infections in a patient with sinusitis by50%. When Mupericin, an antibacterial agent, was formulated in a nasalwash comprising in the 20 mM sodium pyruvate solution and administeredto two (2) patients that suffered from bacterial infections 10 or moretimes annually, the amount of bacterial infections was reduceddramatically.

REFERENCES

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While a number of embodiments of this invention have been represented,it is apparent that the basic construction can be altered to provideother embodiments that utilize the invention without departing from thespirit and scope of the invention. All such modifications and variationsare intended to be included within the scope of the invention as definedin the appended claims rather than the specific embodiments that havebeen presented by way of example.

What is claimed is:
 1. A method for treating a disease state in mammalscharacterized by abnormally low levels of inflammatory agents byup-regulating indigenous in vivo levels of an inflammatory agentcomprising: a. contacting the mammalian cells with an inflammatoryregulator, comprising at least 2.8 mg pyruvate, or a pyruvate precursoror salt thereof, α-keto-isovaleric acid or a precursor thereof andmixtures thereof b. wherein said inflammatory agents are selected fromthe group consisting of elastase, white blood cells, tumor necrosisfactor-α and cytokines selected from the group consisting ofinterleukin-6, interleukin-8, interleukin-10, interleukin-17, andinterleukin-23, and; c. wherein the disease state is selected from thegroup consisting of pulmonary and upper respiratory diseases,Alzheimer's disease, diabetes, and nicotine addiction.
 2. The method ofclaim 1 wherein the pulmonary and upper respiratory disease is selectedfrom the group consisting of infected lungs and infected sinuses,bronchial asthma, acute bronchitis, allergic rhinitis, sinusitis anddiseases caused by organic dust, irritant gases, air pollution andchemicals.
 3. The method of claim 2 wherein the pyruvate and its' saltsare selected from the group consisting of pyruvic acid, lithiumpyruvate, sodium pyruvate, potassium pyruvate, magnesium pyruvate,calcium pyruvate, zinc pyruvate, manganese pyruvate, aluminum pyruvate,ammonium pyruvate, and mixtures thereof.
 4. The method of claim 3wherein the pyruvate precursors are selected from the group consistingof pyruvyl-glycine, pyruvyl-alanine, pyruvyl-leucine, pyruvyl-valine,pyruvyl-isoleucine, pyruvyl-phenylalanine, pyruvamide, salts of pyruvicacid, and mixtures thereof.
 5. The method of claim 4 wherein the dosageof the inflammatory regulator ranges from about 2.8 mg to about 1.0gram.
 6. The method of claim 5 wherein the disease state is an infecteddisease state caused by a bacterial, viral, or fungal infection.
 7. Themethod of claim 6 further comprising contacting the mammalian cells witha therapeutic agent.
 8. The method according to claim 6 wherein thetherapeutic agent is selected from the group consisting ofanti-bacterial agents, anti-viral agents, anti-fungals, anti-tumoragents, anti-histamines, proteins, enzymes, hormones, non-steroidalanti-inflammatory agents, cytokines, steroids, nicotine, and insulin. 9.The method of claim 8 wherein the therapeutic agent is selected from thegroup consisting of insulin.
 10. The method of claim 8 wherein thetherapeutic agent is selected from the group consisting of nicotine. 11.The method of claim 8 wherein the therapeutic agent is a steroidselected from the group consisting of fluticasone, (Flonase®),budesonide (Rhinocort®) beclomethasone, mometasone, flunisolide,triamcinolone and mixtures thereof.
 12. The method of claim 8, whereinthe therapeutic agent is an anti-viral agent selected from the groupconsisting of acyclovir, foscarnet sodium, ribavirin, vidarabine,ganciclovir sodium, Ribavirim, zidovudine, phenol, amantadinehydrochloride, and interferon α-n3, interferon α-2a, and oseltamivir.13. The method of claim 12 wherein the anti-viral agent is selected fromthe group consisting of acyclovir, foscarnet sodium, ribavirin,vidarabine, and ganciclovir sodium.
 14. The method of claim 8, whereinthe therapeutic agent is an anti-biotic agent selected from the groupconsisting of actinomycins, glutarimide antibiotics, sarkomycin,fumagillin, streptonigrin, Mupericin, tenuazonic acid, actinogan,peptinogan, and the anthracyclic antibiotics.
 15. The method of claim 8,wherein the therapeutic agent is an antihistamine selected from thegroup consisting of pseudoephedrine, loratadine, fexofenadine,diphenhydramine, famodidine, ranitidine, citirazine, and other H₁- andH₂-antagonists.
 16. The method of claim 12 wherein the anti-viral agentis present in an amount of from about 0.01% to about 50% by weight. 17.The method of claim 8, wherein the therapeutic agent is a protein forthe treatment of Alzeheimers' disease
 18. The method of claim 17,wherein the protein for the treatment of Alzeheimers' disease isinsulin.
 19. The method of claim 8, wherein the therapeutic agent isadministered prior to delivery of the inflammatory regulator.
 20. Themethod of claim 8, wherein the therapeutic agent is administeredconcomitantly with delivery of the inflammatory regulator.
 21. Themethod of claim 8, wherein the therapeutic agent is administered afterdelivery of the inflammatory regulator.
 22. A method for treating apulmonary and upper respiratory disease state by up-regulatingabnormally low levels of inflammatory agents comprising: a. contactingthe mammalian cells with an inflammatory regulator, comprising at least2.8 mg pyruvate, or a pyruvate precursor or salt thereof,α-keto-isovaleric acid or a precursor thereof and mixtures thereof b.wherein said inflammatory agents are selected from the group consistingof elastase, white blood cells, tumor necrosis factor-α and cytokinesselected from the group consisting of interleukin-6, interleukin-8,interleukin-10, interleukin-17, and interleukin-23, and; c. wherein thedisease state is selected from the group consisting of allergicrhinitis, sinusitis, Alzheimer's disease, diabetes, nicotine addiction,infected lungs and infected sinuses, bronchial asthma, and acutebronchitis.
 23. A method for treating an abnormal pulmonary and upperrespiratory condition by up-regulating abnormally low levels ofinflammatory agents comprising the administration of an inflammatoryregulator, comprising at least 2.8 mg pyruvate, or a pyruvate precursoror salt thereof, α-keto-isovaleric acid or a precursor thereof andmixtures thereof; b. wherein said inflammatory agents are selected fromthe group consisting of elastase, white blood cells, tumor necrosisfactor-α and cytokines selected from the group consisting ofinterleukin-6, interleukin-8, interleukin-10, interleukin-17, andinterleukin-23, and; c. wherein the respiratory condition is selectedfrom the group consisting of sinus irritation and congestion, lungirritation and congestion and snoring.